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Correction of a genetic defect in multipotent germline stem cells using a human artificial chromosome

Gene Therapy volume 15, pages 617–624 (2008)Cite this article

Abstract

Human artificial chromosomes (HACs) have several advantages as gene therapy vectors, including stable episomal maintenance that avoids insertional mutations and the ability to carry large gene inserts including regulatory elements. Multipotent germline stem (mGS) cells have a great potential for gene therapy because they can be generated from an individual's testes, and when reintroduced can contribute to the specialized function of any tissue. As a proof of concept, we herein report the functional restoration of a genetic deficiency in mouse p53−/− mGS cells, using a HAC with a genomic human p53 gene introduced via microcell-mediated chromosome transfer. The p53 phenotypes of gene regulation and radiation sensitivity were complemented by introducing the p53-HAC and the cells differentiated into several different tissue types in vivo and in vitro. Therefore, the combination of using mGS cells with HACs provides a new tool for gene and cell therapies. The next step is to demonstrate functional restoration using animal models for future gene therapy.

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References

  1. Rideout III WM, Hochedlinger K, Kyba M, Daley GQ, Jaenisch R . Correction of a genetic defect by nuclear transplantation and combined cell and gene therapy. Cell 2002; 109: 17–27.

    Article  CAS  Google Scholar 

  2. Kuroiwa Y, Kasinathan P, Matsushita H, Sathiyaselan J, Sullivan EJ, Kakitani M et al. Sequential targeting of the genes encoding immunoglobulin-mu and prion protein in cattle. Nat Genet 2004; 36: 775–780.

    Article  CAS  Google Scholar 

  3. Cossu G, Sampaolesi M . New therapies for muscular dystrophy: cautious optimism. Trends Mol Med 2004; 10: 516–520.

    Article  CAS  Google Scholar 

  4. Odom GL, Gregorevic P, Chamberlain JS . Viral-mediated gene therapy for the muscular dystrophies: successes, limitations and recent advances. Biochim Biophys Acta 2007; 1772: 243–262.

    Article  CAS  Google Scholar 

  5. Pagani F, Buratti E, Stuani C, Bendix R, Dork T, Baralle FE . A new type of mutation causes a splicing defect in ATM. Nat Genet 2002; 30: 426–429.

    Article  CAS  Google Scholar 

  6. O'Connor TP, Crystal RG . Genetic medicines: treatment strategies for hereditary disorders. Nat Rev Genet 2006; 7: 261–276.

    Article  CAS  Google Scholar 

  7. Hacein-Bey-Abina S, Von Kalle C, Schmidt M, McCormack MP, Wulffraat N, Leboulch P et al. LMO2-associated clonal T cell proliferation in two patients after gene therapy for SCID-X1. Science 2003; 302: 415–419.

    Article  CAS  PubMed  Google Scholar 

  8. Kuroiwa Y, Tomizuka K, Shinohara T, Kazuki Y, Yoshida H, Ohguma A et al. Manipulation of human minichromosomes to carry greater than megabase-sized chromosome inserts. Nat Biotechnol 2000; 18: 1086–1090.

    Article  CAS  Google Scholar 

  9. Katoh M, Ayabe F, Norikane S, Okada T, Masumoto H, Horike S et al. Construction of a novel human artificial chromosome vector for gene delivery. Biochem Biophys Res Commun 2004; 321: 280–290.

    Article  CAS  Google Scholar 

  10. Basu J, Willard HF . Artificial and engineered chromosomes: non-integrating vectors for gene therapy. Trends Mol Med 2005; 11: 251–258.

    Article  CAS  Google Scholar 

  11. Dennis C . Cloning: mining the secrets of the egg. Nature 2006; 439: 652–655.

    Article  CAS  Google Scholar 

  12. Hall VJ, Stojkovic P, Stojkovic M . Using therapeutic cloning to fight human disease: a conundrum or reality? Stem Cells 2006; 24: 1628–1637.

    Article  Google Scholar 

  13. Kanatsu-Shinohara M, Inoue K, Lee J, Yoshimoto M, Ogonuki N, Miki H et al. Generation of pluripotent stem cells from neonatal mouse testis. Cell 2004; 119: 1001–1012.

    Article  CAS  PubMed  Google Scholar 

  14. Frengen E, Zhao B, Howe S, Weichenhan D, Osoegawa K, Gjernes E et al. Modular bacterial artificial chromosome vectors for transfer of large inserts into mammalian cells. Genomics 2000; 68: 118–126.

    Article  CAS  Google Scholar 

  15. Horikawa I, Okamoto A, Yokota J, Oshimura M . Genetic heterogeneity of chromosome 11 associated with tumorigenicity in HeLa D98-OR cells. Cancer Genet Cytogenet 1995; 85: 97–100.

    Article  CAS  PubMed  Google Scholar 

  16. Ikeno M, Masumoto H, Okazaki T . Distribution of CENP-B boxes reflected in CREST centromere antigenic sites on long-range alpha-satellite DNA arrays of human chromosome 21. Hum Mol Genet 1994; 3: 1245–1257.

    Article  CAS  PubMed  Google Scholar 

  17. Kawasaki H, Mizuseki K, Nishikawa S, Kaneko S, Kuwana Y, Nakanishi S et al. Induction of midbrain dopaminergic neurons from ES cells by stromal cell-derived inducing activity. Neuron 2000; 28: 31–40.

    Article  CAS  PubMed  Google Scholar 

  18. Bourdon JC, Fernandes K, Murray-Zmijewski F, Liu G, Diot A, Xirodimas DP et al. p53 isoforms can regulate p53 transcriptional activity. Genes Dev 2005; 19: 2122–2137.

    Article  CAS  PubMed  Google Scholar 

  19. Tomizuka K, Yoshida H, Uejima H, Kugoh H, Sato K, Ohguma A et al. Functional expression and germline transmission of a human chromosome fragment in chimaeric mice. Nat Genet 1997; 16: 133–143.

    Article  CAS  PubMed  Google Scholar 

  20. Lin T, Chao C, Saito S, Mazur SJ, Murphy ME, Appella E et al. p53 induces differentiation of mouse embryonic stem cells by suppressing Nanog expression. Nat Cell Biol 2005; 7: 165–171.

    Article  CAS  Google Scholar 

  21. Voet T, Schoenmakers E, Carpentier S, Labaere C, Marynen P . Controlled transgene dosage and PAC-mediated transgenesis in mice using a chromosomal vector. Genomics 2003; 82: 596–605.

    Article  CAS  Google Scholar 

  22. Mejia JE, Willmott A, Levy E, Earnshaw WC, Larin Z . Functional complementation of a genetic deficiency with human artificial chromosomes. Am J Hum Genet 2001; 69: 315–326.

    Article  CAS  PubMed  Google Scholar 

  23. Ayabe F, Katoh M, Inoue T, Kouprina N, Larionov V, Oshimura M . A novel expression system for genomic DNA loci using a human artificial chromosome vector with transformation-associated recombination cloning. J Hum Genet 2005; 50: 592–599.

    Article  CAS  Google Scholar 

  24. Kouprina N, Larionov V . TAR cloning: insights into gene function, long-range haplotypes and genome structure and evolution. Nat Rev Genet 2006; 7: 805–812.

    Article  CAS  Google Scholar 

  25. Copeland NG, Jenkins NA, Court DL . Recombineering: a powerful new tool for mouse functional genomics. Nat Rev Genet 2001; 2: 769–779.

    Article  CAS  Google Scholar 

  26. Wallace HA, Marques-Kranc F, Richardson M, Luna-Crespo F, Sharpe JA, Hughes J et al. Manipulating the mouse genome to engineer precise functional syntenic replacements with human sequence. Cell 2007; 128: 197–209.

    Article  CAS  Google Scholar 

  27. Guan K, Nayernia K, Maier LS, Wagner S, Dressel R, Lee JH et al. Pluripotency of spermatogonial stem cells from adult mouse testis. Nature 2006; 440: 1199–1203.

    Article  CAS  Google Scholar 

  28. Takahashi K, Yamanaka S . Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 2006; 126: 663–676.

    Article  CAS  PubMed  Google Scholar 

  29. Suda T, Katoh M, Hiratsuka M, Takiguchi M, Kazuki Y, Inoue T et al. Heat-regulated production and secretion of insulin from a human artificial chromosome vector. Biochem Biophys Res Commun 2006; 340: 1053–1061.

    Article  CAS  Google Scholar 

  30. Boley SE, Wong VA, French JE, Recio L . p53 heterozygosity alters the mRNA expression of p53 target genes in the bone marrow in response to inhaled benzene. Toxicol Sci 2002; 66: 209–215.

    Article  CAS  Google Scholar 

  31. Otsuki A, Tahimic CG, Tomimatsu N, Katoh M, Chen DJ, Kurimasa A et al. Construction of a novel expression system on a human artificial chromosome. Biochem Biophys Res Commun 2005; 329: 1018–1025.

    Article  CAS  Google Scholar 

  32. Trowell HE, Nagy A, Vissel B, Choo KH . Long-range analyses of the centromeric regions of human chromosomes 13, 14 and 21: identification of a narrow domain containing two key centromeric DNA elements. Hum Mol Genet 1993; 2: 1639–1649.

    Article  CAS  Google Scholar 

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Acknowledgements

We thank K Sato for providing the human p53 cDNA; K Hanaoka, S Tsuji, C Okita, H Yamada, Y Iida and J Nakamura for technical assistance and K Tomizuka, A Kurimasa H Kugoh, T Inoue and M Hiratsuka for valuable discussions. This study was supported in part by a Health and Labor Science Research Grant for Research on the Human Genome, Tissue Engineering from the Ministry of Health, Labour and Welfare, Japan (MO), and the 21st Century COE program from the Ministry of Education, Culture, Sports, Science and Technology of Japan (MO).

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Author notes

  1. J C Barrett

    Present address: 8Current address: Novartis Institutes for BioMedical Research, Cambridge, MA 02139, USA.,

Authors and Affiliations

  1. Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, Japan

    Y Kazuki, H Hoshiya, Y Kai, S Abe, M Takiguchi, M Osaki, S Kawazoe, N Kajitani & M Oshimura

  2. Department of Human Genome Science, Graduate School of Medical Science, Tottori University, Yonago, Tottori, Japan

    M Katoh

  3. Department of Molecular Genetics, Graduate School of Medicine, Kyoto University, Kyoto, Japan

    M Kanatsu-Shinohara & T Shinohara

  4. The Institute of Physical and Chemical Research (RIKEN), Bioresource Center, Ibaraki, Japan

    K Inoue & A Ogura

  5. Division of Functional Genomics, Research Center for Bioscience and Technology, Tottori University, Yonago, Tottori, Japan

    T Yoshino

  6. Department of Genetic Medicine and Regenerative Therapeutics, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, Japan

    Y Shirayoshi

  7. Laboratory of Biosystems and Cancer, National Cancer Institute, Bethesda, MD, USA

    J C Barrett

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Correspondence to M Oshimura.

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Supplementary Information accompanies the paper on Gene Therapy website (http://www.nature.com/gt)

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Kazuki, Y., Hoshiya, H., Kai, Y. et al. Correction of a genetic defect in multipotent germline stem cells using a human artificial chromosome. Gene Ther 15, 617–624 (2008). https://doi.org/10.1038/sj.gt.3303091

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  • DOI: https://doi.org/10.1038/sj.gt.3303091

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